It is well known that aspartase has the ability to convert ammonium fumarate into L-aspartate. Various methods for producing L-aspartate by the enzymatic reaction of aspartase with ammonium fumarate are known. For example, L-aspartic acid can be prepared by cultivating an aspartase-producing microorganism in a nutrient medium containing fumaric acid or fumarate ion. U.S. Pat. No. 3,214,345 discloses the use of E. coli cells to produce L-aspartic acid from ammonium fumarate. Alternatively, L-aspartic acid can be prepared either by reacting resting whole cells containing aspartase with ammonium fumarate or by extracting the enzyme and heating the same with ammonium fumarate. However, these methods are disadvantageous because the resulting L-aspartic acid is contaminated with the enzyme, the microbial cells, nutrient sources, etc. Accordingly, in order to recover L-aspartic acid having high purity, additional steps for removing the enzyme and other contaminants are required. Frequently these methods destroy the enzyme and/or the microorganism so that they can be used only once.
To overcome the above disadvantages, it has been suggested to immobilize the enzyme or enzyme-producing microorganism in or on a support structure. For example, methods of binding or immobilizing enzymes are disclosed in Japanese Patent Publication No. 6870/1970 (binding to anion exchange polysaccharide adsorbent), U.S. Pat. No. 3,672,955 to Stanley (enzymes bound to polyurethane), U.S. Pat. No. 3,975,350 to Hudgin et al. (enzymes entrapped in hydrophilic polymer), and U.S. Pat. Nos. 4,312,946; 3,929,574; or 3,928,138 to Wood et al. (enzymes immobilized in polyurethane foams). Similarly methods of binding or immobilizing microorganisms are disclosed in Japanese Patent Publication No. 17,587/1970 (encapsulated in polyacrylates as polymeric carriers), Russian Patent Publictions Nos. SU 423,976 and SU 451,483 (immobilization of E. coli in polyacrylamide cells), a journal article by Sato, Biochemica et Biophysica Acta, 570,179-186 (1979) (carrageenan as carrier polymer) and pending U.S. patent application Ser. No. 452,579 (Swann et al.), filed Dec. 23, 1982 (immobilized in a polyurethane carrier).
In addition to the preceding references, U.S. Pat. No. 3,391,059 describes a process wherein microorganisms capable of converting maleic acid directly into L-aspartic acid are isolated. In such microorganisms, maleic and/or malonic acid induces formation of enzymes used in the conversion of maleic acid to L-aspartic acid. U.S. Pat. No. 4,013,508 describes conversion of hydrocarbons into L-aspartic acid utilizing two different microorganisms. The first converts hydrocarbons to fumaric acid and the second converts fumaric acid to L-aspartic acid. Maleic acid is produced as a by-product.
Methods for isomerizing maleic acid to fumaric acid are well known. U.S. Pat. No. 2,816,923 describes an isomerization process wherein the maleic acid is heated. It is indicated that isomerization using hydrochloric acid is too expensive and the combination of a halogen (e.g., bromine) plus ultraviolet light is not effective. U.S. Pat. No. 2,955,136 utilizes nitric acid plus a catalyst such as bromine, iodine, hydrochloric acid, hydrobromic acid, potassium thiocyanate or sodium bromide. U.S. Pat. No. 3,332,992 accomplishes the isomerization by use of a bromine-providing catalyst and an oxidizing agent. A suitable oxidizing agent is ammonium persulphate and the bromine-providing catalyst can be ammonium bromide.